One of the hallmarks of CML is the continual evolution of the clinical behavior of the cells throughout the natural history of the disease from the indolent chronic phase to the accelerated and blastic phase in which the patients die of bleeding or infection. Moreover, patients are heterogeneous in phenotype at diagnosis and at each stage of the disease indicating that the genetic changes present at diagnosis vary among patients. Animal experiments also indicate that the bcr/abl gene interacts with other genes, and that once bcr/abl is introduced into cells, chromosomal changes evolve that mirror the changes seen in vivo in cells transduced with the v-abl oncogene. Reproducible chromosomal changes are also seen during the evolution of CML in man. In order to correlate the nature of these genetic changes with responsiveness to therapy, we have devised a novel method of following and characterizing the genetic changes in CML at the chromosomal level following remission induction, the method is sensitive to one leukemic in 10,000 normal cells, depends on FACS sorting based on immunophenotype followed by Fluorescent in-situ hybridization (FISH), and focuses on the following secondary chromosomal abnormalities in CML: duplication of the Philadelphia chromosome [t(9;22)], monosomy 7, trisomy 8, isochromosome 17 and loss of the y chromosome. FACS is used to isolate progenitor cells which are also labeled with BUdR to assess their proliferation. To further investigate genetic instability, we will apply a new technique to identify genetic changes: comparative genomic hybridization (CGH). This technique produces a map of DNA sequence copy numbers as a function of chromosomal location throughout the entire genome and allows the detection of genomic deletions, duplications and amplifications. It is based on the extraction, labeling and comparative hybridization of DNA from CML cells against normal DNA to metaphases from normal cells, and also allows comparison of DNA's from consecutive samples from the same patient. Samples from patients showing disease progression (with and without cytogenetic changes) will be hybridized in early and late stages of their disease and the sequences representing altered loci (amplified or deleted) will be further characterized. These studies should help in identifying the molecular basis for resistance, disease progression and relapse, and thereby help in establishing more effective forms of therapy.